The present invention comprises a process and technique for synthesizing uniformed sized hyaluronic acid (“HA”) particles that are free from oil and surfactant contaminants and useful for a variety of medical and industrial applications.
Hyaluronan (hyaluronic acid; HA), [-b-sodium glucuronic acid-(103)-b-N-acetyl glucosamine-(1®4)-]n, is a polysaccharide made up of repeating disaccharide units of sodium glucuronate and N-acetyl glucosamine linked by b 1-3 and b 1-4 glycosidic bonds. [1] Hyaluronic acid is a known, naturally occurring material, which has many applications in medicine and biology. See, for example, E. A. Balazs U.S. Pat. No. 4,272,522 and publications cited. The isolation and characterization of HA is described by Meyer et al, J. Biol. Chem. 107, 629 (1934); J. Biol. Chem. 114, 689 (1936); Balazs, Fed. Proc. 17, 1086 (1958); Laurent et al; Biochim. Biophys. Acta 42, 476 (1960). The structure of HA was elucidated by Weissman et al, J. Am. Chem. Soc. 76, 1753 (1954) and Meyer, Fed. Proc. 17, 1075 (1958). It should be noted that HA has been used widely used as medical implants for treating different pathological conditions such as eye surgery, osteoarthritis, wound repair and plastic surgery. [2] The highest concentration of HA is found in the soft connective tissue where it is a major component of the extra-cellular matrix, and in the vitreous body of the eye. This jelly-like complex is necessary for transportation of essential nutrients from the bloodstream, via the capillary network, to the living cells of the skin. HA products have been commercialized and widely reported for decades due to their great significance to human body.
The fabrication and applications of the crosslinked HA gel have been reported previously. For example, U.S. Pat. No. 4,636,542 (“the '542 patent”) issued to Hefner, Jr., et al., on Jan. 13, 1987, entitled “Imide Functional Polyphenols; Thermosettable Compositions Containing Same and Cured Products Therefrom,” describes imide functional polyphenols that are prepared by reacting an excess of a phenol with an unsaturated diimide which is prepared by reacting an unsaturated polycycloaliphatic dicarboxylic acid anhydride with a diamine. These materials are useful in the preparation of castings, coatings, laminates and the like.
U.S. Pat. No. 4,500,676 (“the '676 patent”) issued to Balazs, et al., on Feb. 19, 1985, entitled “Hyaluronate Modified Polymeric Articles;” and U.S. Pat. No. 4,487,865 (“the '865 patent”) issued to Balazs, et al., on Dec. 11, 1984, entitled “Polymeric Articles Modified with Hyaluronate;” describe polymeric materials (and articles made therefrom) including polyurethanes, polyesters, polyolefins, polyamides, polysiloxanes, vinylic and acrylic polymers are rendered biocompatible by including with the polymeric material hyaluronic acid or a salt thereof. The hyaluronic acid may be coated onto the surface of the polymeric material, dispersed throughout the body of the polymeric material, or both. The hyaluronic acid on the surface of the polymeric material may optionally be cross-linked. The biocompatible polymeric materials are used in the making of various prosthetic devices including heart valves, intraocular lenses, vascular grafts, pacemaker leads and the like.
U.S. Pat. No. 4,303,676 (“the '4,303,676 patent”) issued to Balazs on Dec. 1, 1981, entitled “Hyaluronate Based Compositions and Cosmetic Formulations Containing Same,” describes a water based, viscoelastic composition for use in cosmetic formulations comprising, (a) a mixture of sodium hyalfractions having different molecular weights, (b) protein which is derived from the natural material from which the hyaluronate is obtained, and (c) water. Also disclosed are cosmetic formulations comprising about 0.05-5.0% of the above composition together with an emollient, a sugar alcohol, a neutral or anionic polysaccharide, a preservative, bacteriostatic and fungistatic substance which does not react with or degrade hyaluronic acid, and water.
U.S. Pat. No. 4,141,976 (“the '976 patent”) issued to Voorhees on Feb. 27, 1979, entitled “Process for alleviating proliferative skin diseases,” describes a process for alleviating proliferative skin diseases such as psoriasis, atopic dermatitis, etc. comprising administering to humans, or domesticated animals, topically and/or systemically a composition comprising a pharmaceutical carrier and at least one active compound selected from the groups, substituted alkyl zanthines, substituted thioxanthines, alone or in combination with a glucocorticoid.
U.S. Pat. No. 3,357,784 (“the '784 patent”) issued to Kasper, et al., on Dec. 12, 1967, entitled “Exposure to Intense Ultraviolet light to Improve Characteristics of Cellulose Fabrics in Divinyl Sulfone and Glyoxal Cross-linking Processes,” describes an advance of treating cellulosic materials with certain unsaturated non-resin forming reactants, whereby enhanced and unexpected stabilization is attained without excessive strength loss.
The entire content of each of: the '542 patent; the '676 patent; the '865 patent; the '4,303,676 patent; the '976 patent; and the '784 patent is hereby incorporated by reference. Generally each of these references describe the formulation and applications of the cross-linking of bulk HA and derivatives with the use of 1,2,3,4-diepoxybutane in alkaline medium at 50° C. (T. C. Laurent, K. Hellsing, and B. Gelotte, Acta Chem. Scand. 18 [1984], No. 1, 274-5). The products obtained by this method are a gel, which substantially swells in water. Divinyl sulfone (“DVS”) has been used previously for cross-linking polysaccharides, especially cellulose, as described in the '784 patent.
Micron-scale HA spheres have been synthesized using conventional water-in-oil emulsion process, spray drying and solvent evaporation. [13] However, it has been difficult to completely remove the oil and surfactant from the HA spheres, and the size and distribution of the HA particles fabricated by spray drying could be hardly controlled as well. The residue of oil and surfactant which are generally prohibited for medical treatment, together with the wide particle size distribution, essentially limited the application of HA as a nano-biomaterial for drug loading, nano-encapsulation and hydrogel architecture designs, such as the fabrication of nanoparticle networks as a drug carrier with two levels hierarchies. Equally important, none of the existing technique can produce monodisperse nanoparticles. Therefore, processes producing uniformed sized HA nanoparticle synthesis without the use of oil and surfactant are highly desirable.
The present invention describes a technique that turns HA into nanometer scaled particulates. Although not wanting to be bound by theory, these nanometer scaled particulates can be used in a variety of medical and industrial applications, including drug delivery vehicles, tissue expander, or tissue engineering scaffolds, whereby creating new medical and industrial applications using state of the art nanotechnology processes and apparatus possible. Additionally, the HA nanoparticles of this invention may be useful to produce drug release device and tissue engineering scaffolds.
It is well recognized that small particle size (on the nanoscale) is useful for carrier-mediated treatment of pathologies that require systemic administration and long-term circulation. The invention of the narrowly distributed surfactant free HA nanoparticle will open a door for the existing commercially available but healthcare-only oriented HA products to the nano-biomaterial for practical medical treatment.